This project focuses on physiologically critical functions of NF-kappaB proteins and their regulators. NF-kappaB is a family of related dimeric transcription factors that serve as primary intracellular mediators of signals that are generated during innate and adaptive immune responses. In addition, dysregulation of NF-kappaB contributes significantly to inflammatory and autoimmune diseases as well as a wide variety of tumors. To identify novel physiologic roles we make use of mouse models engineered to lack components of the NF-kappaB transcription factor family or their regulators. These mutant mice are challenged in vivo in various ways to reveal failed biologic responses. Once a defect is identified this provides the physiologic context in which to dissect out specific cellular and molecular contributions of NF-kappaB proteins or their regulators. NF-kappaB regulators of interest include the classical inhibitory IkappaB proteins, the non-inhibitory IkappaB family protein Bcl-3, as well as several proximal components of signaling pathways that lead to activation of NF-kappaB. We analyze the mutant mice primarily for defects in immune system development, for impaired immune responses to pathogens as well as for failed responses to various stress- or disease-inducing agents. The ultimate goal is to identify critical physiologic processes that depend on NF-kappaB factors and their regulators in health and disease. We aim to identify the relevant cell types involved, the NF-kappaB-activating signaling pathways and the targets of NF-kappaB. We have shown previously that mice deficient in NF-kappaB2 have defects in secondary lymphoid organ architecture and are partially defective in B cell maturation and maintenance. We have demonstrated that the TNF-family member BAFF activates the p52/RelB NF-kappaB dimers in maturing and mature B cells by processing the inhibitory p100 form of NF-kappaB2 to p52. This is the alternative or non-classical pathway for NF-kappaB activation. This pathway is also critical in stromal cells during formation and maintenance of secondary lymphoid architecture, which explains the defects in these organs in NF-kappaB2 deficient mice. This pathway is activated in stromal cells of secondary lymphoid organs by signals transmitted via the lymphotoxin beta receptor, a member of the TNF receptor family. Mice deficient in the NF-kappaB regulatory protein Bcl-3 also display some stromal cell-intrinsic defects in secondary lymphoid architecture, although Bcl-3 does not appear to have a role in the alternative pathway of NF-kappaB activation. We have now generated mice deficient in both NF-kappaB2 and Bcl-3. These compound mutant mice had unexpectedly developed a fatal multi-organ inflammation shortly after birth. This implied partially redundant activities of NF-kappaB 2 and Bcl-3, since singly deficient mice did not develop inflammatory disease. We further determined that this defect was intrinsic to thymic stromal cells. Thymi of compound mutant mice failed to eliminate autoreactive T cells, which in turn resulted in multi-organ inflammatory disease within a few weeks after birth. In another project we were able to demonstrate that NF-kappaB2 contributes to the development of the obesity-induced insulin resistance phenotype in mice, which is a prelude to diabetes type 2 in humans. It has been shown previously that the components of the classical pathway of NF-kappaB activation are required to develop insulin resistance. The present work shows that additional components of the NF-kappaB system also have a role to play, presumably by contributing to a chronic low level inflammatory state which correlates with and may indeed underlie insulin resistance.